Abstract
Eukaryotic initiation factor (eIF) 4B is known to interact with multiple initiation factors, mRNA, rRNA, and poly(A) binding protein (PABP). To gain a better understanding of the function of eIF4B, the two isoforms from Arabidopsis (Arabidopsis thaliana) were expressed and analyzed using biophysical and biochemical methods. Plant eIF4B was found by ultracentrifugation and light scattering analysis to most likely be a monomer with an extended structure. An extended structure would facilitate the multiple interactions of eIF4B with mRNA as well as other initiation factors (eIF4A, eIF4G, PABP, and eIF3). Eight mRNAs, barley (Hordeum vulgare) alpha-amylase mRNA, rabbit beta-hemoglobin mRNA, Arabidopsis heat shock protein 21 (HSP21) mRNA, oat (Avena sativa) globulin, wheat (Triticum aestivum) germin, maize (Zea mays) alcohol dehydrogenase, satellite tobacco necrosis virus RNA, and alfalfa mosaic virus (AMV) 4, were used in wheat germ in vitro translation assays to measure their dependence on eIF4B and eIF4F isoforms. The two Arabidopsis eIF4B isoforms, as well as native and recombinant wheat eIF4B, showed similar responses in the translation assay. AMV RNA 4 and Arabidopsis HSP21 showed only a slight dependence on the presence of eIF4B isoforms, whereas rabbit beta-hemoglobin mRNA and wheat germin mRNA showed modest dependence. Barley alpha-amylase, oat globulin, and satellite tobacco necrosis virus RNA displayed the strongest dependence on eIF4B. These results suggest that eIF4B has some effects on mRNA discrimination during initiation of translation. Barley alpha-amylase, oat globulin, and rabbit beta-hemoglobin mRNA showed the highest activity with eIF4F, whereas Arabidopsis HSP21 and AMV RNA 4 used both eIF4F and eIF(iso)4F equally well. These results suggest that differential or optimal translation of mRNAs may require initiation complexes composed of specific isoforms of initiation factor gene products. Thus, individual mRNAs or classes of mRNAs may respond to the relative abundance of a particular initiation factor(s), which in turn may affect the amount of protein translated. It is likely that optimal multifactor initiation complexes exist that allow for optimal translation of mRNAs under a variety of cellular conditions.
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